The present invention relates to an automatic DNA purification apparatus and more particularly, to an apparatus for isolating and purifying DNA automatically from various liquids which contain DNA, such as cell culture media.
As is known, the process for DNA isolation and purification from DNA containing solutions, such as cell culture media, is prerequisite procedure for gene amplification, nucleotide sequence analysis, gene recombination. At present, several conventional methods for isolation and purification of DNA from various biological samples, such as blood, biological tissue, animal cell, plant tissue, E.coli, virus, agarose gel and the like, have been publicly known.
For example, the boiling method (Holms, D. S. and M. Quigley, 1981, Anal. Biochem. 114:193) and the alkaline lysis method (Birnboim, H. C. and J. Doly, 1979, Nucleic Acids Res. 7:1513) have been reported as methods for isolation of plasmid DNA from E.coli and bacterial transformants.
Also, the plasmid DNA purification method which employs CsCl density gradient centrifugation process to obtain high purity DNA, has been reported. However, such method takes a lot of time and in addition, comprises a lot of steps, such as detergent-mediated lysis step, protease treatment step, phenol extraction and ethanol precipitation step. Therefore, such method is not suitable to be employed as a commercial process for the production of high purity DNA in a large scale.
In addition, it has also been reported that the DNA purification method, wherein DNA is adsorbed on the surface of silica or glass in the presence of chaotropic salts, such as NaI or NaClO4 and thereby, isolated from various extracts (Marko, M. A. et al. 1982, A procedure for the large scale isolation of highly purified DNA using alkaline extraction and binding to glass powder. Anal. Biochem. 121:382-387; Vogestein. B. et al. 1979. Preparative and analytical purification of DNA from agarose. Proc. Natl. Acad. Sci. USA. 76:615-619). Such DNA adsorption method in the presence of chaotropic salts, is a general method which isolates and purifies chromosomal DNA or plasmid DNA from cell lysate (Buffone, G. J., et al. 1991. Clin. Chem. 37., 1945).
Such adsorption process is proceeded by using compressed glass fiber or silica particle suspension which contain activated surface on which DNA is adsorbed in guanidine hydrochloride buffer solution with a pH of 4 to 5 or in 4M sodium iodide solution with a pH of 7.5 to 8.
Recently, DNA purification apparatus which can obtain DNA at a time from a large number of biological samples, is required for genome study, blood test, plant seed inspection, agricultural test, microbiological environmental test.
By the way, as mentioned above, although DNA isolation and purification process is a prerequisite step for nucleotide sequence analysis, gene amplification or gene cloning process, the conventional hand-operated DNA purification process is time-consuming and very laborious process and thus, the amount of DNA obtained therefrom varied remarkably depends on operator""s mastery.
In order to overcome the above problems, several automatic purification apparatuses have been developed up to now. However, the number of DNA solutions which can be treated at a time in the apparatuses thus developed, is limited. Moreover, DNA cannot be obtained in a large amount within a short time because it is impossible to supply DNA containing solutions continuously and sequentially in such apparatus.
In addition, the conventional DNA purification apparatus is not cost-effective due to the disposable injection tips which should be used for the injection of various solutions and, could not be automated completely on account of filtration and adsorption process.
Therefore, in this technical field, the development of automatic DNA purification apparatus which can isolate and purify DNA simultaneously from a large number of liquids which contain DNA through automatic processes, and which do not use disposable injection tips by employing injection needles which can be used repeatedly, has been desired.
In order to produce such automatic DNA purification apparatus, the injection apparatus which can select liquids from various samples automatically and sequentially and which can control precisely the quantitative injection of the selected liquids, has been desired.
However, the rotary type multi-channel valve has been developed up to present, has some drawbacks, for example, the leakage of liquid in contacting point between each flow passages and the formation of liquid droplet in terminal of outlet, which obstruct seriously accurate quantitative liquid injection. Thus, such type multi-channel valve cannot be employed in an automatic DNA purification apparatus which should control precisely quantitative and selective injection of a large number of liquid in due sequence.
Therefore, the development new type multi-channel quantitative control valve which can minimize the leakage of liquid, and which can accurately control the quantitative injection of liquid by suppressing the formation of liquid droplet in the terminal of flow passage by means of pulse-mode liquid feeding, has been required for the production of high efficient automatic DNA purification apparatus.
In addition, an injection apparatus which can feed various kinds of liquids continuously and in due sequence, is prerequisite for the production of the automatic DNA purification apparatus which can isolate and purify DNA simultaneously from various kinds of liquids which contain DNA through automatic processes.
Therefore, the objection of the present invention is to provide an apparatus which can isolate and purify DNA in large scale within a short time simultaneously from various liquids which contain DNA.
The above object of the present invention is achieved by providing an automatic DNA purification apparatus, which comprises:
liquid containers (10) which supply various liquid reagents and liquids which contain DNA;
numerous liquid flow passages (21) wherein various kind of liquids stored in said liquid container (10) are conveyed;
control valves (22, 24, 25) which control the selection and transfer of various liquids;
numerous syringes (23) which intake and discharge said various liquids quantitatively;
numerous injection needles (26) which are connected with said numerous syringes (23) respectively through numerous liquid flow passages;
transportation mean (30) for moving said injection needles and/or multi-well plate toward up-down and/or right-left directions;
vacuum block (41) wherein liquids contained in said multi-well plate are filtered in vacuo;
multi-well plate rack (31) which transports said multi-well plates toward up-down directions;
numerous multi-well plate blocks (40) wherein multi-well plates which have been transported through said multi-well plate rack, are mounted;
process control unit(not depicted) which regulates the operation of said control valves for selection and transfer of said various liquids, the operation of said syringes for quantitative intake and discharge of said liquids, the transfer of said injection needles connected with said syringes, the sequential transportation of said multi-well plates arranged in said multi-well plate rack, the sequential operation of shaking block and said vacuum block, according to a computer program.
In addition, the automatic DNA purification apparatus of the present invention may further comprises a washing block (60) which washes said injection needles and/or a shaking block (50) which shake said multi-well plate.
The operation of this apparatus is controlled by said process control unit which instructs, regulates or intermits the sequential proceeding of all operations according to the computer program installed therein in advance. The mechanical operations of this apparatus driven by electric motors, are regulated by electric signal generated from said process control unit. The operations of said control valves and the transfer of liquids are regulated by air pressure generated from air compressor and/or air pump(not depicted) according to electric signals of said process control unit.
Said process control unit is composed of conventional computer equipped with memory units, central processing units, input-output units and the likes disclosed to a public. The computer programs which regulate all procedures for isolation and purification of DNA, are installed in said process control unit, and which may be programmed in common process control software commercialized.